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Biomedical subjects

G Kroemer

Publications and source records attributed to G Kroemer.

At least 109 records · Page 6Linked to original sources
PubMed2000

Permeabilization of the mitochondrial inner membrane during apoptosis: impact of the adenine nucleotide translocator.

Mitochondrial membrane permeabilization can be a rate limiting step of apoptotic as well as necrotic cell death. Permeabilization of the outer mitochondrial membrane (OM) and/or inner membrane (IM) is, at least in part, mediated by the permeability transition pore complex (PTPC). The PTPC is formed in the IM/OM contact site and contains the two most abundant IM and OM proteins, adenine nucleotide translocator (ANT, in the IM) and voltage-dependent anion channel (VDAC, in the OM), the matrix protein cyclophilin D, which can interact with ANT, as well as apoptosis-regulatory proteins from the Bax/Bcl-2 family. Here we discuss that ANT has two opposite functions. On the one hand, ANT is a vital, specific antiporter which accounts for the exchange of ATP and ADP on IM. On the other hand, ANT can form a non-specific pore, as this has been shown by electrophysiological characterization of purified ANT reconstituted into synthetic lipid bilayers or by measuring the permeabilization of proteoliposomes containing ANT. Pore formation by ANT is induced by a variety of different agents (e.g. Ca(2+), atractyloside, thiol oxidation, the pro-apoptotic HIV-1 protein Vpr, etc.) and is enhanced by Bax and inhibited by Bcl-2, as well as by ADP. In isolated mitochondria, pore formation by ANT leads to an increase in IM permeability to solutes up to 1500 Da, swelling of the mitochondrial matrix, and OM permeabilization, presumably due to physical rupture of OM. Although alternative mechanisms of mitochondrial membrane permeabilization may exist, ANT emerges as a major player in the regulation of cell death. Cell Death and Differentiation (2000) 7, 1146 - 1154

Animals↗
PubMed2000

Apoptosis and apoptosis-associated perturbations of peripheral blood lymphocytes during HIV infection: comparison between AIDS patients and asymptomatic long-term non-progressors.

This study was designed to compare the degree of lymphocyte apoptosis and Fas-Fas ligand (FasL) expression in AIDS patients and long-term non-progressors (LTNPs) and correlate these parameters with apoptosis-associated perturbations in lymphocyte function. LTNPs had a lower frequency of apoptotic CD4+ and CD8+ T cells compared with subjects with AIDS. This correlated with a lower frequency of cells expressing Fas and FasL. The frequency of selected lymphocyte populations exhibiting a disrupted mitochondrial transmembrane potential (DeltaPsim) and increased superoxide generation was lower in LTNPs than in patients with AIDS; these abnormalities were associated with lower levels of caspase-1 activation in LTNPs. The results indicate a significantly reduced level of apoptosis and apoptosis-associated parameters in LTNPs than in patients developing AIDS. Based on these findings, a crucial role for mitochondria can be predicted in the process of lymphocyte apoptosis during the evolution of AIDS.

Acquired Immunodeficiency Syndrome↗
PubMed2000

GD3 ganglioside directly targets mitochondria in a bcl-2-controlled fashion.

Lipid and glycolipid diffusible mediators are involved in the intracellular progression and amplification of apoptotic signals. GD3 ganglioside is rapidly synthesized from accumulated ceramide after the clustering of death-inducing receptors and triggers apoptosis. Here we show that GD3 induces dissipation of DeltaPsim and swelling of isolated mitochondria, which results in the mitochondrial release of cytochrome c, apoptosis inducing factor, and caspase 9. Soluble factors released from GD3-treated mitochondria are sufficient to trigger DNA fragmentation in isolated nuclei. All these effects can be blocked by cyclosporin A, suggesting that GD3 is acting at the level of the permeability transition pore complex. We found that endogenous GD3 accumulates within mitochondria of cells undergoing apoptosis after ceramide exposure. Accordingly, suppression of GD3 synthase (ST8) expression in intact cells substantially prevents ceramide-induced DeltaPsim dissipation, indicating that endogenously synthesized GD3 induces mitochondrial changes in vivo. Finally, enforced expression of bcl-2 significantly prevents GD3-induced mitochondrial changes, caspase 9 activation, and apoptosis. These results show that mitochondria are a key destination for apoptogenic GD3 ganglioside along the lipid pathway to programmed cell death and indicate that relevant GD3 targets are under bcl-2 control.

Animals↗
PubMed2000

Mitochondrial control of cell death induced by HIV-1-encoded proteins.

In most examples of physiological or pathological cell death, mitochondrial membrane permeabilization (MMP) constitutes an early critical event of the lethal process. Signs of MMP that precede nuclear apoptosis include the translocation of cytochrome c and apoptosis-inducing factor (AIF) from mitochondria to an extra-mitochondrial localization, as well as the dissipation of the mitochondrial transmembrane potential. MMP also occurs in HIV-1-induced apoptosis. Different HIV-1 encoded proteins (Env, Vpr, Tat, PR) can directly or indirectly trigger MMP, thereby causing cell death. The gp120/gp41 Env complex constitutes an example for an indirect MMP inducer. Env expressed on the plasma membrane of HIV-1 infected (or Env-transfected) cells mediates cell fusion with CD4/CXCR4-expressing uninfected cells. After a cell type-dependent latency period, syncytia then undergo MMP and apoptosis. Vpr exemplifies a direct MMP inducer. Vpr binds to the adenine nucleotide translocator (ANT), a mitochondrial inner membrane protein which also interacts with apoptosis-regulatory proteins from the Bcl-2/Bax family. Binding of Vpr to ANT favors formation of a non-specific pore leading to MMP. The structural motifs of the Vpr protein involved in MMP are conserved among most pathogenic HIV-1 isolates and determine the cytotoxic effect of Vpr. These data suggest the possibility that viruses employ multiple strategies to regulate host cell apoptosis by targeting mitochondria.

Animals↗
PubMed2000

Mitochondrio-nuclear translocation of AIF in apoptosis and necrosis.

Apoptosis inducing factor (AIF) is a novel apoptotic effector protein that induces chromatin condensation and large-scale ( approximately 50 kbp) DNA fragmentation when added to purified nuclei in vitro. Confocal and electron microscopy reveal that, in normal cells, AIF is strictly confined to mitochondria and thus colocalizes with heat shock protein 60 (hsp60). On induction of apoptosis by staurosporin, c-Myc, etoposide, or ceramide, AIF (but not hsp60) translocates to the nucleus. This suggests that only the outer mitochondrial membrane (which retains AIF in the intermembrane space) but not the inner membrane (which retains hsp60 in the matrix) becomes protein permeable. The mitochondrio-nuclear redistribution of AIF is prevented by a Bcl-2 protein specifically targeted to mitochondrial membranes. The pan-caspase inhibitor Z-VAD. fmk does not prevent the staurosporin-induced translocation of AIF, although it does inhibit oligonucleosomal DNA fragmentation and arrests chromatin condensation at an early stage. ATP depletion is sufficient to cause AIF translocation to the nucleus, and this phenomenon is accelerated by the apoptosis inducer staurosporin. However, in conditions in which both glycolytic and respiratory ATP generation is inhibited, cells fail to manifest any sign of chromatin condensation and advanced DNA fragmentation, thus manifesting a 'necrotic' phenotype. Both in the presence of Z-VAD. fmk and in conditions of ATP depletion, AIF translocation correlates with the appearance of large-scale DNA fragmentation. Altogether, these data are compatible with the hypothesis that AIF is a caspase-independent mitochondrial death effector responsible for partial chromatinolysis.

Adenosine Triphosphate↗
PubMed2000

Apoptosis and cell cycle: distinct checkpoints with overlapping upstream control.

The question as to whether apoptosis (programmed cell death) is controlled by one or few checkpoints is still unresolved. A growing body of evidence suggests that (one of) the decisive event(s) of cell death consists in the permeabilization of mitochondrial membranes. Indeed, multiple pro-apopotic signal transduction pathways converge on the proteins of the Bcl-2/Bax family which, in concert with the so-called permeability transition pore complex (PTPC), regulate mitochondrial membrane barrier function. Mitochondrial permeabilization causes the release of soluble intermembrane proteins, some of which are involved in the activation of apoptotic proteases and nucleases. Thus, the putative checkpoint determining the death/life decision is clearly different from the known checkpoints of cell cycle progression. Prominent oncogenes (e.g., c-Myc, Ras, Raf, Bcl-2) and tumor suppressor genes (e.g., p53, Bax) have been shown to modulate apoptosis via a direct or indirect effect on mitochondrial membranes. All these oncoproteins and tumor suppressor proteins may simultaneously influence the cell cycle and the propensity to undergo apoptosis. Several cell cycle regulatory proteins (e.g., cyclins, cdk, etc.) can induce or inhibit apoptosis via yet unknown pathways.

Animals↗
PubMed1999

The novel retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphtalene carboxylic acid can trigger apoptosis through a mitochondrial pathway independent of the nucleus.

The novel retinoid 6-[3-(1-adamantyl)-4-hydroxyphenyl]-2-naphtalene carboxylic acid (AHPN/CD437), a retinoic acid receptor (RAR)gamma activator, has been found to inhibit the growth and to induce apoptosis of a wide variety of malignant cell types including solid tumors and various leukemias. Interestingly, CD437 is able to induce apoptosis in some all-trans-retinoic acid (ATRA)-resistant models. In a number of experimental systems, the early apoptotic stage that precedes nuclear chromatinolysis consists in mitochondrial alterations, including a disruption of the inner mitochondrial transmembrane potential (delta(psi)m) mediated by the mitochondrial permeability transition (MPT). Similarly CD437 causes RPMI 8226, a human myeloma cell line, to undergo a rapid delta(psi)m disruption that precedes other apoptotic alterations such as the generation of reactive oxygen species and DNA fragmentation. The same sequence of events is observed during the CD437-induced apoptosis in L363, a RARgamma-negative human myeloma cell line, as well as RPMI 8226 cytoplasts (anucleate cells). Indeed, RPMI 8226 cells and cytoplasts manifest a similar degree in delta(psi)m loss, phosphatidylserine exposure, and caspase activation in response to CD437, which indicates that nuclear effects cannot account for the apoptogenic potential of CD437. The mitochondrial release of cytochrome c, the activation of caspases as well as nuclear signs of CD437-induced apoptosis are fully prevented by the MPT inhibitory compound cyclosporin A. Purified mitochondria can be directly induced to undergo MPT with CD437 but not with ATRA. In a cell-free in vitro system consisting of exposing mitochondrial supernatants to isolated nuclei, only supernatants from CD437-treated mitochondria provoke chromatin condensation, whereas supernatants from mitochondria treated with ATRA, or with the combination of CD437 and cyclosporin A, remain inactive. In conclusion, these results suggest that the rapid execution of CD437-induced apoptosis is a nucleus-independent (and probably RARgamma-independent) phenomenon involving mitochondria and MPT.

Antineoplastic Agents↗
PubMed1999

Cytochrome c.

Explore the source record for details and available documents.

Apoptosis↗
PubMed1999

Arsenite induces apoptosis via a direct effect on the mitochondrial permeability transition pore.

The molecular mode of action of arsenic, a therapeutic agent employed in the treatment of acute promyelocytic leukemia, has been elusive. Here we provide evidence that arsenic compounds may act on mitochondria to induce apoptosis. Arsenite induces apoptosis accompanied by a loss of the mitochondrial transmembrane potential (Delta Psim). Inhibition of caspases prevents the arsenite-induced nuclear DNA loss, but has no effect on the Delta Psim dissipation and cytolysis induced by arsenite. In contrast, Bcl-2 expression induced by gene transfer prevents all hallmarks of arsenite-induced cell death, including the Delta Psim collapse. PK11195, a ligand of the mitochondrial benzodiazepine receptor, neutralizes this Bcl-2 effect. Mitochondria are required in a cell-free system to mediate arsenite-induced nuclear apoptosis. Arsenite causes the release of an apoptosis-inducing factor (AIF) from the mitochondrial intermembrane space. This effect is prevented by the permeability transition (PT) pore inhibitor cyclosporin A, as well as by Bcl-2, which is known to function as an endogenous PT pore antagonist. Arsenite also opens the purified, reconstituted PT pore in vitro in a cyclosporin A- and Bcl-2-inhibitible fashion. Altogether these data suggest that arsenite can induce apoptosis via a direct effect on the mitochondrial PT pore.

Apoptosis↗
PubMed1999

Plasma membrane potential in thymocyte apoptosis.

Apoptosis is accompanied by major changes in ion compartmentalization and transmembrane potentials. Thymocyte apoptosis is characterized by an early dissipation of the mitochondrial transmembrane potential, with transient mitochondrial swelling and a subsequent loss of plasma membrane potential (DeltaP sip) related to the loss of cytosolic K+, cellular shrinkage, and DNA fragmentation. Thus, a gross perturbation of DeltaPsip occurs at the postmitochondrial stage of apoptosis. Unexpectedly, we found that blockade of plasma membrane K+ channels by tetrapentylammonium (TPA), which leads to a DeltaP sip collapse, can prevent the thymocyte apoptosis induced by exposure to the glucocorticoid receptor agonist dexamethasone, the topoisomerase inhibitor etoposide, gamma-irradiation, or ceramide. The TPA-mediated protective effect extends to all features of apoptosis, including dissipation of the mitochondrial transmembrane potential, loss of cytosolic K+, phosphatidylserine exposure on the cell surface, chromatin condensation, as well as caspase and endonuclease activation. In strict contrast, TPA is an ineffective inhibitor when cell death is induced by the potassium ionophore valinomycin, the specific mitochondrial benzodiazepine ligand PK11195, or by primary caspase activation by Fas/CD95 cross-linking. These results underline the importance of K+ channels for the regulation of some but not all pathways leading to thymocyte apoptosis.

Animals↗
PubMed1999

Lonidamine triggers apoptosis via a direct, Bcl-2-inhibited effect on the mitochondrial permeability transition pore.

The molecular mode of action of lonidamine, a therapeutic agent employed in cancer chemotherapy, has been elusive. Here we provide evidence that lonidamine (LND) acts on mitochondria to induce apoptosis. LND provokes a disruption of the mitochondrial transmembrane potential which precedes signs of nuclear apoptosis and cytolysis. The mitochondrial and cytocidal effects of LND are not prevented by inhibitors of caspases or of mRNA or protein synthesis. However, they are prevented by transfection-enforced overexpression of Bcl-2, an oncoprotein which inhibits apoptosis by stabilizing the mitochondrial membrane barrier function. Accordingly, the cell death-inducing effect of LND is amplified by simultaneous addition of PK11195, an isoquinoline ligand of the peripheral benzodiazepine receptor which antagonizes the cytoprotective effect of Bcl-2. When added to isolated nuclei, LND fails to provoke DNA degradation unless mitochondria are added simultaneously. In isolated mitochondria, LND causes the dissipation of the mitochondrial inner transmembrane potential and the release of apoptogenic factors capable of inducing nuclear apoptosis in vitro. Thus the mitochondrion is the subcellular target of LND. All effects of LND on isolated mitochondria are counteracted by cyclosporin A, an inhibitor of the mitochondrial PT pore. We therefore tested the effect of LND on the purified PT pore reconstituted into liposomes. LND permeabilizes liposomal membranes containing the PT pore. This effect is prevented by addition of recombinant Bcl-2 protein but not by a mutant Bcl-2 protein that has lost its apoptosis-inhibitory function. Altogether these data indicate that LND represents a novel type of anti-cancer agent which induces apoptosis via a direct effect on the mitochondrial PT pore.

Amino Acid Chloromethyl Ketones↗
PubMed1999

Molecular characterization of mitochondrial apoptosis-inducing factor.

Mitochondria play a key part in the regulation of apoptosis (cell death). Their intermembrane space contains several proteins that are liberated through the outer membrane in order to participate in the degradation phase of apoptosis. Here we report the identification and cloning of an apoptosis-inducing factor, AIF, which is sufficient to induce apoptosis of isolated nuclei. AIF is a flavoprotein of relative molecular mass 57,000 which shares homology with the bacterial oxidoreductases; it is normally confined to mitochondria but translocates to the nucleus when apoptosis is induced. Recombinant AIF causes chromatin condensation in isolated nuclei and large-scale fragmentation of DNA. It induces purified mitochondria to release the apoptogenic proteins cytochrome c and caspase-9. Microinjection of AIF into the cytoplasm of intact cells induces condensation of chromatin, dissipation of the mitochondrial transmembrane potential, and exposure of phosphatidylserine in the plasma membrane. None of these effects is prevented by the wide-ranging caspase inhibitor known as Z-VAD.fmk. Overexpression of Bcl-2, which controls the opening of mitochondrial permeability transition pores, prevents the release of AIF from the mitochondrion but does not affect its apoptogenic activity. These results indicate that AIF is a mitochondrial effector of apoptotic cell death.

Amino Acid Sequence↗
PubMed1999

Mitochondrial release of caspase-2 and -9 during the apoptotic process.

The barrier function of mitochondrial membranes is perturbed early during the apoptotic process. Here we show that the mitochondria contain a caspase-like enzymatic activity cleaving the caspase substrate Z-VAD.afc, in addition to three biological activities previously suggested to participate in the apoptotic process: (a) cytochrome c; (b) an apoptosis-inducing factor (AIF) which causes isolated nuclei to undergo apoptosis in vitro; and (c) a DNAse activity. All of these factors, which are biochemically distinct, are released upon opening of the permeability transition (PT) pore in a coordinate, Bcl-2-inhibitable fashion. Caspase inhibitors fully neutralize the Z-VAD.afc-cleaving activity, have a limited effect on the AIF activity, and have no effect at all on the DNase activities. Purification of proteins reacting with the biotinylated caspase substrate Z-VAD, immunodetection, and immunodepletion experiments reveal the presence of procaspase-2 and -9 in mitochondria. Upon induction of PT pore opening, these procaspases are released from purified mitochondria and become activated. Similarly, upon induction of apoptosis, both procaspases redistribute from the mitochondrion to the cytosol and are processed to generate enzymatically active caspases. This redistribution is inhibited by Bcl-2. Recombinant caspase-2 and -9 suffice to provoke full-blown apoptosis upon microinjection into cells. Altogether, these data suggest that caspase-2 and -9 zymogens are essentially localized in mitochondria and that the disruption of the outer mitochondrial membrane occurring early during apoptosis may be critical for their subcellular redistribution and activation.

Animals↗
PubMed1999

Pathophysiology of mitochondrial cell death control.

Mitochondria have been recently recognized to play a major role in the control of apoptosis or programmed cell death. Permeabilization of mitochondrial membranes, a decisive feature of early cell death, is regulated by members of the Bcl-2 family which interact with the permeability transition pore complex (PTPC). Thus, the cytoprotective oncoprotein Bcl-2 stabilizes the mitochondrial membrane barrier function, whereas the tumor suppressor protein Bax permeabilizes mitochondrial membranes. The regulation of membrane permeabilization is intertwined with that of the bioenergetic and redox functions of mitochondria. The implications of alterations in the composition of the PTPC and in mitochondrial function for the pathophysiology of cancer (reduced apoptosis) and neurodegeneration (enhanced apoptosis) are discussed.

Animals↗
PubMed1999

Palmitate induces apoptosis via a direct effect on mitochondria.

The fatty acid palmitate can induce apoptosis. Here we show that the palmitate-induced dissipation of the mitochondrial transmembrane potential (Delta Psi m), which precedes nuclear apoptosis, is not prevented by inhibitors of mRNA synthesis, protein synthesis, caspases, or pro-apoptotic ceramide signaling. However, the mitochondrial and nuclear effects of palmitate are inhibited by overexpression of anti-apoptotic proto-oncogene product Bcl-2 and exacerbated by 2-bromo-palmitate as well as by carnitine. The cytoprotective actions of Bcl-2, respectively, is not antagonized by etomoxir, an inhibitor of carnitine palmitoyl transferase 1 (CPT1), suggesting that the recently described physical interaction between CPT1 and Bcl-2 is irrelevant to Bcl-2-mediated inhibition of palmitate-induce apoptosis. When added to purified mitochondria, palmitate causes the release of soluble factors capable of stimulating the apoptosis of isolated nuclei in a cell-free system. Mitochondria purified from Bcl-2 over-expressing cells are protected against the palmitate-stimulated release of such factors. These data suggest that palmitate causes apoptosis via a direct effect on mitochondria.

Journal Article↗